Genomics Application Notes

Materials and Methods The data was generated using the materials and methods described in the field installation guide.

Results and Discussion 25ng and 100ng Universal Human Reference (UHR) RNA (Agilent Technologies) was used as input for the New England Biolabs NEBNext Ultra Directional RNA Library Prep Kit for Illumina on the Beckman Coulter Biomek FX P automated liquid handler. UHR RNA was assayed on the Agilent Bioanalyzer 2100 using an RNA6000 Pico kit (PN# 5067-1513) prior to the run to ensure high sample quality (data not shown). The purified adapter ligated library was amplified to 15 cycles per instruction of the New England Biolabs NEBNext Ultra Directional RNA Library Prep Kit for Illumina protocol using the Bio-Rad S1000 Thermal Cycler (Bio-Rad., Inc). The quality and size distribution of the library was analyzed using the Agilent DNA High Sensitivity kit (PN# 5067- 4626) and Agilent Bioanalyzer 2100. Representative Bioanalyzer 2100 results are shown in Figure 2. The library concentrations were measured using KAPA SYBR Fast Universal 2X qPCR Master Mix library quantification kits (PN# KK4824, KAPA Biosystems), and are summarized in Figure 3.

Figure 2. 25ng UHR RNA (left) and 100ng UHR RNA (right) NEBNext Ultra Directional RNA libraries for Illumina sequencing.

AAG-161TCH07.14-A